ASSESSMENT OF POTENTIAL SARS-CoV-2 VIRUS N GENE INTEGRATION INTO HUMAN GENOME REVEALS NO SIGNIFICANT IMPACT ON RT-qPCR COVID-19 DIAGNOSTIC TESTING

ABSTRACT

The SARS Coronavirus 2 (SARS-CoV-2) pandemic presents new scientific and scale-up challenges for diagnostic capabilities worldwide. The gold standard diagnostic for SARS-CoV-2 infection is a reverse transcription/quantitative PCR (RT-qPCR) which targets the viral genome, an assay that has now been performed on millions of patient specimens worldwide regardless of symptomatic status. Recently Zhang et al. suggested the possibility that the SARS-CoV-2 N gene could integrate into host cell DNA through the action of the LINE-1 retrotransposon, a mobile element that is potentially active in human somatic cells, thereby calling into question the veracity of N-gene based RT-qPCR for detection of SARS-CoV-2 infection. Accordingly, we assessed the potential impact of these purported integration events on nasal swab specimens tested at our clinical laboratory. Using an N-gene based RT-qPCR assay, we tested 768 arbitrarily selected specimens and identified 2 samples which resulted in a positive detection of viral sequence in the absence of reverse transcriptase, a necessary but not sufficient signal consistent with possible integration of the SARS-CoV-2 N gene into the host genome. Regardless of possible viral N gene integration into the genome, in this small subset of samples, all patients were still positive for SARS-CoV-2 infection, as indicated by a much lower Ct value for reactions performed in the presence of reverse transcriptase (RT) versus reactions performed without RT. Moreover, one of the two positives observed in the absence of RT also tested positive when using primers targeting ORF1ab, a gene closer to the 5 end of the genome. These data are inconsistent with the N gene integration hypothesis suggested by the studies by Zhang et al., and importantly, our results suggest little to no practical impact of possible SARS-CoV-2 genome integration events on RT-qPCR testing. COMPETING INTEREST STATEMENTThe authors of this study are employees of the Pandemic Response Lab (PRL)/ReOpen Diagnostics, a private company performing SARS-CoV-2 RT-qPCR based testing, an area of interest of this study.